The primary objectives of this proposal are two folds: test the hypothesis that the diverse biological responses elicited by the multifunctional cytokine TGF-( are mediated by a combination of the canonical pathway of T(RI(Alk5)-Smad2/3 and other pathways such as those involving Alk1- Smad1/5;and test the hypothesis that the specificity and intensity of TGF-( signaling in different cellular context are also predetermined by the basal level of Smad3 due to the presence of a regulatory apparatus consisting of Axin-GSK3( that mediates the turnover of Smad3 in the absence of TGF-( signal. As shown in the section of Preliminary Studies, we have generated substantial amount of preliminary data to support the physiological significance of the topics and the feasibility of the research plan. Thus, Aim 1 will determine the function and mechanism by which TGF-( elicits specific biological responses, such as inhibition of B lymphocyte proliferation and stimulation of cell migration, via the non-canonical bone morphogenetic factor associated receptor I/Smad1/5 signaling pathway. Validation of this hypothesis would challenge the current paradigm in the field that Smad1/5 function primarily as effectors for the BMP signaling pathway and only very rarely mediate TGF-( signal as in endothelial cells. Aim 2 will determine the functional significance of and mechanism underlying the turnover of basal level of Smad3 by the Axin-GSK3( complex. We will study the biochemical and biological nature of phosphorylation of specific residues on Smad3 by the GSK3( kinase. Since APC and CKI have been defined as integral parts of the (-catenin destruction complex, we will also investigate whether these two proteins are involved in Smad3 turnover. While the paradigm for the primary TGF-( signaling pathway has been well established, much more needs to be learned on the contributions of non-canonical pathways to the mediation of diverse biological responses in specific cellular contexts. Accomplishment of these specific aims will lead to a better understanding on the actions and mechanisms of this important factor in cell homeostasis, development, and disease processes. As a multifunctional cytokine, TGF-( is involved in the regulation of many physiological and pathological processes. A better understanding of the mechanisms underlying the actions of TGF-( could help the development of novel therapeutics for many human diseases, including cancer and fibrosis.